Please note: this site relies heavily on the use of javascript. Without a javascript-enabled browser, this site will not function correctly. Please enable javascript and reload the page, or switch to a different browser.
2044  structures 2517  species 23  interactions 22248  sequences 596  architectures

Family: Trypsin (PF00089)

Summary: Trypsin

Pfam includes annotations and additional family information from a range of different sources. These sources can be accessed via the tabs below.

This is the Wikipedia entry entitled "Trypsin". More...

Trypsin Edit Wikipedia article

Trypsin
Identifiers
EC number 3.4.21.4
CAS number 9002-07-7
Databases
IntEnz IntEnz view
BRENDA BRENDA entry
ExPASy NiceZyme view
KEGG KEGG entry
MetaCyc metabolic pathway
PRIAM profile
PDB structures RCSB PDB PDBe PDBsum
Gene Ontology AmiGO / EGO
Trypsin
1UTN.png
Identifiers
Symbol Trypsin
Pfam PF00089
InterPro IPR001254
SMART SM00020
PROSITE PDOC00124
MEROPS S1
SCOP 1c2g
SUPERFAMILY 1c2g
CDD cd00190

Trypsin (EC 3.4.21.4) is a serine protease from the PA clan superfamily, found in the digestive system of many vertebrates, where it hydrolyses proteins.[2][3] Trypsin is produced in the pancreas as the inactive proenzyme trypsinogen. Trypsin cleaves peptide chains mainly at the carboxyl side of the amino acids lysine or arginine, except when either is followed by proline. It is used for numerous biotechnological processes. The process is commonly referred to as trypsin proteolysis or trypsinisation, and proteins that have been digested/treated with trypsin are said to have been trypsinized.

Function

In the duodenum, trypsin catalyzes the hydrolysis of peptide bonds, breaking down proteins into smaller peptides. The peptide products are then further hydrolyzed into amino acids via other proteases, rendering them available for absorption into the blood stream. Tryptic digestion is a necessary step in protein absorption as proteins are generally too large to be absorbed through the lining of the small intestine.

Trypsin is produced in the pancreas, in the form of the inactive zymogen trypsinogen. When the pancreas is stimulated by cholecystokinin, it is then secreted into the first part of the small intestine (the duodenum) via the pancreatic duct. Once in the small intestine, the enzyme enteropeptidase activates it into trypsin by proteolytic cleavage. Auto catalysis can happen with trypsin with trypsinogen as the substrate. This activation mechanism is common for most serine proteases, and serves to prevent autodegradation of the pancreas.

Mechanism

The enzymatic mechanism is similar to that of other serine proteases. These enzymes contain a catalytic triad consisting of histidine-57, aspartate-102, and serine-195.[4] These three residues form a charge relay that serves to make the active site serine nucleophilic. This is achieved by modifying the electrostatic environment of the serine. The enzymatic reaction that trypsin catalyzes is thermodynamically favorable but requires significant activation energy (it is "kinetically unfavorable"). In addition, trypsin contains an "oxyanion hole" formed by the backbone amide hydrogen atoms of Gly-193 and Ser-195, which serves to stabilize the developing negative charge on the carbonyl oxygen atom of the cleaved amides.

The aspartate residue (Asp 189) located in the catalytic pocket (S1) of trypsin is responsible for attracting and stabilizing positively charged lysine and/or arginine, and is, thus, responsible for the specificity of the enzyme. This means that trypsin predominantly cleaves proteins at the carboxyl side (or "C-terminal side") of the amino acids lysine and arginine except when either is bound to a C-terminal proline.,[5] although large-scale mass spectrometry data suggest cleavage occurs even with proline.[6] Trypsin is considered an endopeptidase, i.e., the cleavage occurs within the polypeptide chain rather than at the terminal amino acids located at the ends of polypeptides.

Properties

Trypsin has an optimal operating pH of about 7.5-8.5 and optimal operating temperature of about 37.1°C.[5]

As a protein trypsin has various molecular weights depending on the source. For example, a molecular weight of 23.3 kDa is reported for trypsin from bovine and porcine sources.


The activity of trypsin is not affected by the enzyme inhibitor tosyl phenylalanyl chloromethyl ketone, TPCK, which deactivates chymotrypsin. This is important because, in some applications, like mass spectrometry, the specificity of cleavage is important.

Trypsin should be stored at very cold temperatures (between −20°C and −80°C) to prevent autolysis, which may also be impeded by storage of trypsin at pH 3 or by using trypsin modified by reductive methylation. When the pH is adjusted back to pH 8, activity returns.

Isozymes

The following human genes encode proteins with trypsin enzymatic activity:

protease, serine, 1 (trypsin 1)
Identifiers
Symbol PRSS1
Alt. symbols TRY1
Entrez 5644
HUGO 9475
OMIM 276000
RefSeq NM_002769
UniProt P07477
Other data
Locus Chr. 7 q32-qter
protease, serine, 2 (trypsin 2)
Identifiers
Symbol PRSS2
Alt. symbols TRYP2
Entrez 5645
HUGO 9483
OMIM 601564
RefSeq NM_002770
UniProt P07478
Other data
Locus Chr. 7 q35
protease, serine, 3 (mesotrypsin)
Identifiers
Symbol PRSS3
Alt. symbols PRSS4
Entrez 5646
HUGO 9486
OMIM 613578
RefSeq NM_002771
UniProt P35030
Other data
Locus Chr. 9 p13

Other isoforms of trypsin may also be found in other organisms.

Clinical significance

Activation of trypsin from proteolytic cleavage of trypsinogen in the pancreas can lead to a series of events that cause pancreatic self-digestion, resulting in pancreatitis. One consequence of the autosomal recessive disease cystic fibrosis is a deficiency in transport of trypsin and other digestive enzymes from the pancreas. This leads to the disorder termed meconium ileus. This disorder involves intestinal obstruction (ileus) due to overly thick meconium, which is normally broken down by trypsin and other proteases, then passed in feces.[7]

Applications

Trypsin is available in high quantity in pancreases, and can be purified rather easily. Hence it has been used widely in various biotechnological processes.

In a tissue culture lab, trypsin is used to re-suspend cells adherent to the cell culture dish wall during the process of harvesting cells.[8] Some cell types have a tendency to "stick" - or adhere - to the sides and bottom of a dish when cultivated in vitro. Trypsin is used to cleave proteins bonding the cultured cells to the dish, so that the cells can be suspended in fresh solution and transferred to fresh dishes.

Trypsin can also be used to dissociate dissected cells (for example, prior to cell fixing and sorting).

Trypsin can be used to break down casein in breast milk. If trypsin is added to a solution of milk powder, the breakdown of casein will cause the milk to become translucent. The rate of reaction can be measured by using the amount of time it takes for the milk to turn translucent.

Trypsin is commonly used in biological research during proteomics experiments to digest proteins into peptides for mass spectrometry analysis, e.g. in-gel digestion. Trypsin is particularly suited for this, since it has a very well defined specificity, as it hydrolyzes only the peptide bonds in which the carbonyl group is contributed either by an Arg or Lys residue.

Trypsin can also be used to dissolve blood clots in its microbial form and treat inflammation in its pancreatic form.

In food

Commercial protease preparations usually consist of a mixture of various protease enzymes that often includes trypsin. These preparations are widely utilized in food processing:[9]

  • as a baking enzyme to improve the workability of dough;
  • in the extraction of seasonings and flavourings from vegetable or animal proteins and in the manufacture of sauces;
  • to control aroma formation in cheese and milk products;
  • to improve the texture of fish products;
  • to tenderize meat;
  • during cold stabilization of beer;
  • in the production of hypoallergenic food where proteases break down specific allergenic proteins into nonallergenic peptides. For example, proteases are used to produce hypoallergenic baby food from cow’s milk thereby diminishing the risk of babies developing milk allergies.

Trypsin inhibitor

Main article: Trypsin inhibitor

In order to prevent the action of active trypsin in the pancreas which can be highly damaging, inhibitors such as BPTI and SPINK1 in the pancreas and α1-antitrypsin in the serum are present as part of the defense against its inappropriate activation. Any trypsin prematurely formed from the inactive trypsinogen would be bound by the inhibitor. The protein-protein interaction between trypsin and its inhibitors is one of the tightest found, and trypsin is bound by some of its pancreatic inhibitors essentially irreversibly.[10] In contrast with nearly all known protein assemblies, some complexes of trypsin bound by its inhibitors do not readily dissociate after treatment with 8M urea.[11]

See also

Trypsinization PA clan of proteases

References

  1. ^ PDB 1UTN; Leiros HK, Brandsdal BO, Andersen OA, Os V, Leiros I, Helland R, Otlewski J, Willassen NP, Smalås AO (April 2004). "Trypsin specificity as elucidated by LIE calculations, X-ray structures, and association constant measurements". Protein Sci. 13 (4): 1056–70. doi:10.1110/ps.03498604. PMC 2280040. PMID 15044735. 
  2. ^ Rawlings ND, Barrett AJ (1994). "Families of serine peptidases". Meth. Enzymol. Methods in Enzymology 244: 19–61. doi:10.1016/0076-6879(94)44004-2. ISBN 978-0-12-182145-6. PMID 7845208. 
  3. ^ The German physiologist Wilhelm Kühne (1837-1900) discovered trypsin in 1876. See: W. Kühne (1877) "Über das Trypsin (Enzym des Pankreas)", Verhandlungen des naturhistorisch-medicinischen Vereins zu Heidelberg, new series, vol. 1, no. 3, pages 194-198.
  4. ^ Polgár L (October 2005). "The catalytic triad of serine peptidases". Cell. Mol. Life Sci. 62 (19–20): 2161–72. doi:10.1007/s00018-005-5160-x. PMID 16003488. 
  5. ^ a b "Sequencing Grade Modified Trypsin". www.promega.com. 2007-04-01. Retrieved 2009-02-08. 
  6. ^ Rodriguez J, Gupta N, Smith RD, Pevzner PA (2008). "Does trypsin cut before proline?". J. Proteome Res. 7 (1): 300–305. doi:10.1021/pr0705035. PMID 18067249. 
  7. ^ Noone PG, Zhou Z, Silverman LM, Jowell PS, Knowles MR, Cohn JA (December 2001). "Cystic fibrosis gene mutations and pancreatitis risk: relation to epithelial ion transport and trypsin inhibitor gene mutations". Gastroenterology 121 (6): 1310–9. doi:10.1053/gast.2001.29673. PMID 11729110. 
  8. ^ "Trypsin-EDTA (0.25%)". Stem Cell Technologies. Retrieved 2012-02-23. 
  9. ^ "Protease - GMO Database". GMO Compass. European Union. 2010-07-10. Retrieved 2012-01-01. 
  10. ^ Voet & Voet (1995). Biochemisty (2nd ed.). John Wiley & Sons. pp. 396–400. ISBN 0-471-58651-X. 
  11. ^ N. Levilliers, M. Péron, B. Arrio, J. Pudles (October 1970). "On the mechanism of action of proteolyticinhibitors: IV. Effect of 8murea on the stability of trypsin in trypsin-lnhibitor complexes". Archives of Biochemistry and Biophysics 140 (2): 474–483. PMID 5528741. 

External links

.

This page is based on a Wikipedia article. The text is available under the Creative Commons Attribution/Share-Alike License.

This tab holds the annotation information that is stored in the Pfam database. As we move to using Wikipedia as our main source of annotation, the contents of this tab will be gradually replaced by the Wikipedia tab.

Trypsin Provide feedback

No Pfam abstract.

Literature references

  1. Rawlings ND, Barrett AJ; , Meth Enzymol 1994;244:19-61.: Families of Serine Peptidases PUBMED:7845208 EPMC:7845208

  2. Sprang S, Standing T, Fletterick RJ, Stroud RM, Finer-Moore J, Xuong NH, Hamlin R, Rutter WJ, Craik CS; , Science 1987;237:905-909.: The Three Dimensional Structure of Asnl02 Trypsin: Role of Aspl02 in Serine Protease Catalysis PUBMED:3112942 EPMC:3112942


Internal database links

External database links

This tab holds annotation information from the InterPro database.

InterPro entry IPR001254

In the MEROPS database peptidases and peptidase homologues are grouped into clans and families. Clans are groups of families for which there is evidence of common ancestry based on a common structural fold:

  • Each clan is identified with two letters, the first representing the catalytic type of the families included in the clan (with the letter 'P' being used for a clan containing families of more than one of the catalytic types serine, threonine and cysteine). Some families cannot yet be assigned to clans, and when a formal assignment is required, such a family is described as belonging to clan A-, C-, M-, N-, S-, T- or U-, according to the catalytic type. Some clans are divided into subclans because there is evidence of a very ancient divergence within the clan, for example MA(E), the gluzincins, and MA(M), the metzincins.
  • Peptidase families are grouped by their catalytic type, the first character representing the catalytic type: A, aspartic; C, cysteine; G, glutamic acid; M, metallo; N, asparagine; S, serine; T, threonine; and U, unknown. The serine, threonine and cysteine peptidases utilise the amino acid as a nucleophile and form an acyl intermediate - these peptidases can also readily act as transferases. In the case of aspartic, glutamic and metallopeptidases, the nucleophile is an activated water molecule. In the case of the asparagine endopeptidases, the nucleophile is asparagine and all are self-processing endopeptidases.

In many instances the structural protein fold that characterises the clan or family may have lost its catalytic activity, yet retain its function in protein recognition and binding.

Proteolytic enzymes that exploit serine in their catalytic activity are ubiquitous, being found in viruses, bacteria and eukaryotes [PUBMED:7845208]. They include a wide range of peptidase activity, including exopeptidase, endopeptidase, oligopeptidase and omega-peptidase activity. Many families of serine protease have been identified, these being grouped into clans on the basis of structural similarity and other functional evidence [PUBMED:7845208]. Structures are known for members of the clans and the structures indicate that some appear to be totally unrelated, suggesting different evolutionary origins for the serine peptidases [PUBMED:7845208].

Not withstanding their different evolutionary origins, there are similarities in the reaction mechanisms of several peptidases. Chymotrypsin, subtilisin and carboxypeptidase C have a catalytic triad of serine, aspartate and histidine in common: serine acts as a nucleophile, aspartate as an electrophile, and histidine as a base [PUBMED:7845208]. The geometric orientations of the catalytic residues are similar between families, despite different protein folds [PUBMED:7845208]. The linear arrangements of the catalytic residues commonly reflect clan relationships. For example the catalytic triad in the chymotrypsin clan (PA) is ordered HDS, but is ordered DHS in the subtilisin clan (SB) and SDH in the carboxypeptidase clan (SC) [PUBMED:7845208, PUBMED:8439290].

This group of serine proteases belong to the MEROPS peptidase family S1 (chymotrypsin family, clan PA(S))and to peptidase family S6 (Hap serine peptidases).

The chymotrypsin family is almost totally confined to animals, although trypsin-like enzymes are found in actinomycetes of the genera Streptomyces and Saccharopolyspora, and in the fungus Fusarium oxysporum [PUBMED:7845208]. The enzymes are inherently secreted, being synthesised with a signal peptide that targets them to the secretory pathway. Animal enzymes are either secreted directly, packaged into vesicles for regulated secretion, or are retained in leukocyte granules [PUBMED:7845208].

The Hap family, 'Haemophilus adhesion and penetration', are proteins that play a role in the interaction with human epithelial cells. The serine protease activity is localized at the N-terminal domain, whereas the binding domain is in the C-terminal region.

Gene Ontology

The mapping between Pfam and Gene Ontology is provided by InterPro. If you use this data please cite InterPro.

Domain organisation

Below is a listing of the unique domain organisations or architectures in which this domain is found. More...

Loading domain graphics...

Pfam Clan

Alignments

We store a range of different sequence alignments for families. As well as the seed alignment from which the family is built, we provide the full alignment, generated by searching the sequence database using the family HMM. We also generate alignments using four representative proteomes (RP) sets, the NCBI sequence database, and our metagenomics sequence database. More...

View options

We make a range of alignments for each Pfam-A family. You can see a description of each above. You can view these alignments in various ways but please note that some types of alignment are never generated while others may not be available for all families, most commonly because the alignments are too large to handle.

  Seed
(71)
Full
(22248)
Representative proteomes NCBI
(28849)
Meta
(4196)
RP15
(2699)
RP35
(3841)
RP55
(6981)
RP75
(9786)
Jalview View  View  View  View  View  View  View  View 
HTML View    View  View         
PP/heatmap 1   View  View         
Pfam viewer View  View             

1Cannot generate PP/Heatmap alignments for seeds; no PP data available

Key: ✓ available, x not generated, not available.

Format an alignment

  Seed
(71)
Full
(22248)
Representative proteomes NCBI
(28849)
Meta
(4196)
RP15
(2699)
RP35
(3841)
RP55
(6981)
RP75
(9786)
Alignment:
Format:
Order:
Sequence:
Gaps:
Download/view:

Download options

We make all of our alignments available in Stockholm format. You can download them here as raw, plain text files or as gzip-compressed files.

  Seed
(71)
Full
(22248)
Representative proteomes NCBI
(28849)
Meta
(4196)
RP15
(2699)
RP35
(3841)
RP55
(6981)
RP75
(9786)
Raw Stockholm Download   Download   Download   Download   Download   Download   Download   Download  
Gzipped Download   Download   Download   Download   Download   Download   Download   Download  

You can also download a FASTA format file containing the full-length sequences for all sequences in the full alignment.

External links

MyHits provides a collection of tools to handle multiple sequence alignments. For example, one can refine a seed alignment (sequence addition or removal, re-alignment or manual edition) and then search databases for remote homologs using HMMER3.

HMM logo

HMM logos is one way of visualising profile HMMs. Logos provide a quick overview of the properties of an HMM in a graphical form. You can see a more detailed description of HMM logos and find out how you can interpret them here. More...

Trees

This page displays the phylogenetic tree for this family's seed alignment. We use FastTree to calculate neighbour join trees with a local bootstrap based on 100 resamples (shown next to the tree nodes). FastTree calculates approximately-maximum-likelihood phylogenetic trees from our seed alignment.

Note: You can also download the data file for the tree.

Curation and family details

This section shows the detailed information about the Pfam family. You can see the definitions of many of the terms in this section in the glossary and a fuller explanation of the scoring system that we use in the scores section of the help pages.

Curation View help on the curation process

Seed source: SCOP and Prosite
Previous IDs: trypsin;
Type: Domain
Author: Lutfiyya LL, Sonnhammer ELL
Number in seed: 71
Number in full: 22248
Average length of the domain: 206.20 aa
Average identity of full alignment: 23 %
Average coverage of the sequence by the domain: 59.84 %

HMM information View help on HMM parameters

HMM build commands:
build method: hmmbuild -o /dev/null HMM SEED
search method: hmmsearch -Z 23193494 -E 1000 --cpu 4 HMM pfamseq
Model details:
Parameter Sequence Domain
Gathering cut-off 20.6 20.6
Trusted cut-off 20.6 20.6
Noise cut-off 20.5 20.5
Model length: 220
Family (HMM) version: 21
Download: download the raw HMM for this family

Species distribution

Sunburst controls

Show

This visualisation provides a simple graphical representation of the distribution of this family across species. You can find the original interactive tree in the adjacent tab. More...

Loading sunburst data...

Tree controls

Hide

The tree shows the occurrence of this domain across different species. More...

Loading...

Please note: for large trees this can take some time. While the tree is loading, you can safely switch away from this tab but if you browse away from the family page entirely, the tree will not be loaded.

Interactions

There are 23 interactions for this family. More...

Trypsin VWA WAP Squash LRR_1 PDZ SSI Sema TIL Propep_M14 Staphylokinase Antistasin Ecotin Pacifastin_I Kazal_2 V-set Hepsin-SRCR EGF Kunitz_BPTI PAN_1 Kazal_1 Serpin Sushi

Structures

For those sequences which have a structure in the Protein DataBank, we use the mapping between UniProt, PDB and Pfam coordinate systems from the PDBe group, to allow us to map Pfam domains onto UniProt sequences and three-dimensional protein structures. The table below shows the structures on which the Trypsin domain has been found. There are 2044 instances of this domain found in the PDB. Note that there may be multiple copies of the domain in a single PDB structure, since many structures contain multiple copies of the same protein seqence.

Loading structure mapping...